Electrostatic and structural properties of complexes involving plasmid DNA and cationic lipids commonly used for gene delivery.
The present study is aimed to characterize the interactions between plasmid DNA and cationic, large unilamellar vesicles, 110+/-20nm in size, composed of lipids commonly used for transfections including DOTAP/DOPE (mole ratio 1/1), DOTAP/DOPC (mole ratio 1/1), 100% DOTAP, or DC-CHOL/DOPE (mole ratio 1/1). [


BACKGROUND
DOTAP, N-(1-(2,3-dioleoyloxy)propyl)-N,N,N-trimethylammonium chloride; DOPE, 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine; DOPC, 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine; DC-CHOL, 3 beta-[N-(N',N'-dimethylaminoethane)carbamoyl] cholesterol]. A novel approach of combining Gouy-Chapman calculations and fluorescence measurements of the pH at the surface of lipid assemblies by the fluorophore 4-heptadecyl-7-hydroxycoumarin showed that electrostatic parameters played a key role in the instantaneous formation of the DNA-lipid complexes upon addition of different amounts of plasmid DNA to cationic liposomes in 20 mM Hepes buffer (pH 7.4). Addition of large amounts of plasmid DNA leads to neutralization of 60% of the protonated DC-CHOL in DC-CHOL/DOPE (1/1) assemblies and 80% of the DOTAP in lipid assemblies. The characterization of these electrostatic parameters of the complexes suggests better and closer surrounding of plasmid DNA by lipids when DOPE is present. Time-dependent static light-scattering measurements monitored the formation of complexes and also showed that these complexes were highly unstable with respect to size at DNA/cationic lipid molar ratios between 0.2 and 0.8.
Zuidam
N J
NJ
Department of Biochemistry, The Hebrew University-Hadassah Medical School, Jerusalem, Israel.
Barenholz
Y
Y
D002412:Cations; D004274:DNA, Recombinant / Q000737:chemistry*; D018014:Gene Transfer Techniques; D008027:Light; D008055:Lipids / Q000737:chemistry*; D010957:Plasmids / Q000235:genetics*; D012542:Scattering, Radiation; D013050:Spectrometry, Fluorescence; D055672:Static Electricity
D002412:Cations; D004274:DNA, Recombinant; D008055:Lipids